title: One-step peptide backbone dissociations in negative-ion free radical initiated peptide sequencing mass spectrometry.
abstract: Peptide dissociation behavior in TEMPO (2,2,6,6-tetramethylpiperidine-1-oxyl)-based FRIPS (free radical initiated peptide sequencing) mass spectrometry was analyzed in both positive- and negative-ion modes for a number of peptides including angiotensin II, kinetensin, glycoprotein IIb fragment (296-306), des-Pro(2)-bradykinin, and ubiquitin tryptic fragment (43-48). In the positive mode, the ·Bz-C(O)-peptide radical species was produced exclusively at the initial collisional activation of o-TEMPO-Bz-C(O)-peptides, and two consecutive applications of collisional activation were needed to observe peptide backbone fragments. In contrast, in the negative-ion mode, a single application of collisional activation to o-TEMPO-Bz-C(O)-peptides produced extensive peptide backbone fragmentations as well as ·Bz-C(O)-peptide radical species. This result indicates that the duty cycle in the TEMPO-based FRIPS mass spectrometry can be reduced by one-half in the negative-ion mode. In addition, the fragment ions observed in the negative-ion experiments were mainly of the a-, c-, x-, and z-types, indicating that radical-driven tandem mass spectrometry was mainly responsible for the TEMPO-based FRIPS even with a single application of collisional activation. Furthermore, the survival fraction analysis of o-TEMPO-Bz-C(O)-peptides was made as a function of the applied normalized collision energy (NCE). This helped us to better understand the differences in FRIPS behavior between the positive- and negative-ion modes in terms of dissociation energetics. The duty-cycle improvement made in the present study provides a cornerstone for future research aiming to achieve a single-step FRIPS in the positive-ion mode.
authors.affiliation: Department of Chemistry, Sogang University, Seoul 121-742, Korea.
authors.forename: Jihye
authors.initials: J
authors.lastname: Lee
authors.forename: Hyeyeon
authors.initials: H
authors.lastname: Park
authors.forename: Hyuksu
authors.initials: H
authors.lastname: Kwon
authors.forename: Gyemin
authors.initials: G
authors.lastname: Kwon
authors.forename: Aeran
authors.initials: A
authors.lastname: Jeon
authors.forename: Hugh I
authors.initials: HI
authors.lastname: Kim
authors.forename: Bong June
authors.initials: BJ
authors.lastname: Sung
authors.forename: Bongjin
authors.initials: B
authors.lastname: Moon
authors.forename: Han Bin
authors.initials: HB
authors.lastname: Oh
mesh_terms: D000595:Amino Acid Sequence; D003497:Cyclic N-Oxides / Q000737:chemistry; D005609:Free Radicals / Q000737:chemistry; D008958:Models, Molecular; D010455:Peptides / Q000737:chemistry*; D011487:Protein Conformation; D020539:Sequence Analysis, Protein / Q000379:methods*; D053719:Tandem Mass Spectrometry / Q000379:methods*
chemical_list: D003497:Cyclic N-Oxides; D005609:Free Radicals; D010455:Peptides; C003959:TEMPO